This proposal seeks to explain how the lac repressor recognizes the unique base sequence of the lac operator. Alone, this protein binds specifically and very tightly to the lac operator, but a variety of galactosides induce it to no longer bind. These inducers must alter features of the protein which are essential for its DNA binding activity. Conformational changes promoted by inducer binding have been detected. However, the methods used do not specifically reveal what is happening at the DNA binding site of the repressor. By attaching fluorescent probes to this site, the relevant conformational changes might be directly observed. Recently, changes in DNA structure have been detected after repessor binding. The possibility that the repressor undergoes yet another conformational change at this time needs to be investigated. We have selectively attached fluorescent probes to tyrosine residues in the lac repressor. The location of these probes will be investigated by limited tryptic digestion of the modified repressor. We have also selectively attached fluorescent probes to other sites on the repressor. These probes will be used to detect conformational changes when the repressor binds either inducer or DNA. The kinetics of any change will be investigated by stopped-flow fluorimetric methods.